Toxin proteins such as diphtheria toxin (DT) typically are made up of several functional domains, which may include a toxin (i.e., killing) domain, an internalization domain (e.g., the DT translocation enhancing region (TER)), and a targeting domain to control recognition of and binding to target cells. DT is a single chain of 535 amino acids (Greenfield et al. (1983) Proc. Natl. Acad. Sci. USA 80:6853-6857), which upon mild trypsinization and reduction in vitro breaks into an A chain and a B chain (Collier et al. (1971) J. Biol. Chem. 246:1496-503; and Moskaug et al. (1989) J. Biol. Chem. 264:15709-15713). The B chain contains the targeting domain and the TER, which facilitates translocation of the A chain into the cytoplasm. Once in the cytoplasm, the toxin domain within the A chain catalyzes ADP-ribosylation of a translationally modified histidine residue (diphthamide) on elongation factor-2, leading to the arrest of protein synthesis and subsequent cell death (Collier et al., ADP Ribosylation Reactions: Biology and Medicine, Academic Press, Inc., New York, p. 573 (1982)). Other toxins (e.g., ricin) have similar A and B chain structures, while toxins such as the Pseudomonas exotoxin have similar domains but in a single chain structure.
Fusion toxins can be therapeutically useful in pathological conditions such as cancer, and particularly in types of cancer (e.g., certain brain cancers) that are unresponsive to treatment by chemotherapy and radiation. Fusion toxins are chimeric polypeptides that typically contain a toxin protein or a toxin domain from a toxin protein, and a targeting domain from a heterologous protein (Kreitman (1999) Curr. Opin. Immunol. 11:570-578; and Oldfield and Youle (1998) Curr. Top. Microbiol. Immunol. 234:97-114). Fusion toxins may incorporate a portion of a toxin protein or an entire toxin protein (see, for example, Pastan and FitzGerald (1989) J. Biol. Chem. 264:15157-15160). Fusion toxins that contain an entire toxin molecule, however, typically result in non-specific killing mediated by binding to non-target cells. Fusion toxins that contain only the toxin (killing) domain of a toxin protein, while much more specific, are much less toxic because they lack the translocation enhancing region that facilitates entry of the toxin domain into target cells.
Malignancies of the central nervous system are the third leading cause of cancer-related deaths among adolescents and adults from 15 to 34 years of age (Davis et al. (1998) J. Neurosurg. 88:1-10). Patients with such malignancies typically have a two-year survival rate of less than 20% (Thompson (1995) Science 267:1414). Although the anatomy of brain tumors would especially lend them to intratumoral therapy with agents such as fusion toxins, therapeutic approaches to treating such tumors are complicated by the fact that there has been no known tumor-specific marker that can be targeted in the majority of patients (McKeever (1998) J. Histochem. Cytochem. 46:585-594).